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Translation of rat intestinal RNA yields two alkaline phosphatases.

ABSTRACT: After translation of total rat intestinal RNA, immunoprecipitation using monospecific antiserum against rat intestinal alkaline phosphatase yielded two polypeptides in the adult duodenum and jejunum (molecular masses 62 and 65 kDa). Immunoprecipitation of both bands was blocked by a single purified alkaline phosphatase. In the adult ileum and in the entire small intestine of suckling pups, only the 62 kDa translation product was found. After fat feeding, translated alkaline phosphatase increased by an amount proportionate to the increase in enzyme activity previously seen in the serum. A small fraction of nascent alkaline phosphatase was translocated into microsomal vesicles, producing peptides of 65 and 69 kDa. Tunicamycin-treated membranes demonstrated a different signal peptide for each translation product. N-Terminal sequencing of the translation products showed leucine residues at similar positions, but overlap with the mature protein sequence was not demonstrated. On the basis of these data, we propose the presence of two mRNAs encoding alkaline phosphatase in the rat intestine.

SUBMITTER: Sussman NL

PROVIDER: S-EPMC1146608 | biostudies-other | 1986 Mar

REPOSITORIES: biostudies-other

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Project description:Mouse serum alkaline phosphatase (ALP) is frequently measured and interpreted in mammalian bone research. However, little is known about the circulating ALPs in mice and their relation to human ALP isozymes and isoforms. Mouse ALP was extracted from liver, kidney, intestine, and bone from vertebra, femur and calvaria tissues. Serum from mixed strains of wild-type (WT) mice and from individual ALP knockout strains were investigated, i.e., Alpl(-/-) (a.k.a. Akp2 encoding tissue-nonspecific ALP or TNALP), Akp3(-/-) (encoding duodenum-specific intestinal ALP or dIALP), and Alpi(-/-) (a.k.a. Akp6 encoding global intestinal ALP or gIALP). The ALP isozymes and isoforms were identified by various techniques and quantified by high-performance liquid chromatography. Results from the WT and knockout mouse models revealed identical bone-specific ALP isoforms (B/I, B1, and B2) as found in human serum, but in addition mouse serum contains the B1x isoform only detected earlier in patients with chronic kidney disease and in human bone tissue. The two murine intestinal isozymes, dIALP and gIALP, were also identified in mouse serum. All four bone-specific ALP isoforms (B/I, B1x, B1, and B2) were identified in mouse bones, in good correspondence with those found in human bones. All mouse tissues, except liver and colon, contained significant ALP activities. This is a notable difference as human liver contains vast amounts of ALP. Histochemical staining, Northern and Western blot analyses confirmed undetectable ALP expression in liver tissue. ALP activity staining showed some positive staining in the bile canaliculi for BALB/c and FVB/N WT mice, but not in C57Bl/6 and ICR mice. Taken together, while the main source of ALP in human serum originates from bone and liver, and a small fraction from intestine (<5%), mouse serum consists mostly of bone ALP, including all four isoforms, B/I, B1x, B1, and B2, and two intestinal ALP isozymes dIALP and gIALP. We suggest that the genetic nomenclature for the Alpl gene in mice (i.e., ALP liver) should be reconsidered since murine liver has undetectable amounts of ALP activity. These findings should pave the way for the development of user-friendly assays measuring circulating bone-specific ALP in mouse models used in bone and mineral research.

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Translation of rat intestinal RNA yields two alkaline phosphatases.

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